The pharmacology and toxicology of general anesthetics are remarkably incomplete for such a widely used and medically important class of drugs that are administered to increasingly older and sicker patients. Knowledge of the mechanisms of anesthetic action is insufficient to explain how any anesthetic produces amnesia, unconsciousness or immobilization (with increasing doses), the cardinal features of general anesthesia. Anesthetics have potent and specific effects on synaptic transmission, including both presynaptic actions on the release of neurotransmitters and postsynaptic actions on receptors. The principal objective of this research proposal is to understand the presynaptic mechanisms of anesthetic effects on neurotransmitter release by experimentally isolating these effects from their better understood postsynaptic actions. Presynaptic actions could be involved in therapeutic effects (unconsciousness, amnesia, immobility) and/or their toxic effects (neurotoxicity, respiratory depression, cardiovascular depression) of anesthetics. Understanding synaptic mechanisms of anesthetics is essential for development of anesthetics with improved side-effect profiles and for optimization of current anesthetic techniques in high-risk patients. We have shown that general anesthetics inhibit glutamate release by presynaptic mechanisms and that these effects are transmitter-specific and involve region-specific inhibition of specific Na+ channel subtypes. We now propose to focus on the region- and transmitter-specific actions and Na+ channel blocking mechanisms of volatile anesthetics in order to more fully understand their presynaptic actions. Our central hypothesis is that general anesthetics affect neurotransmitter release by synapse-specific mechanisms due to effects on presynaptic ion channels. We will test this hypothesis using an integrative and collaborative multidisciplinary approach by the following Specific Aims: Aim 1-Determine the mechanisms by which volatile anesthetics differentially affect neurotransmitter release from isolated nerve terminals to test the hypothesis that they have synapse-specific effects on transmitter release due to differences in presynaptic mechanisms; Aim 2-Determine the neurotransmitter-specific effects and mechanisms of volatile anesthetics on exocytosis in intact neurons to test the hypothesis that they differentially inhibit synaptic vesicle exocytosis by neurotransmitter-specific and ion channel-dependent mechanisms; and Aim 3-Determine the mechanisms and regulation of volatile anesthetic effects on voltage- gated Na+ channels to test the hypothesis that they inhibit Na+ channel subtypes by state-dependent mechanisms. Complementary approaches include analysis of anesthetic effects on transmitter release from intact nerve terminals, synaptic vesicle exocytosis from single cultured hippocampal neurons, and biophysical properties of specific Na+ channel subtypes. Such studies are essential to a molecular understanding of presynaptic anesthetic mechanisms and the balance between desirable and potentially toxic anesthetic effects on excitatory and inhibitory synaptic transmission. PUBLIC HEALTH RELEVANCE: The molecular and cellular mechanisms by which anesthetics produce amnesia, unconsciousness and immobilization, the cardinal features of general anesthesia, are unknown despite their critical role in the modern pharmacopoeia. We have shown that inhaled anesthetics inhibit neurotransmitter release from nerve terminals by inhibition of Na+ channels involving mechanisms that vary with the specific anesthetic, neurotransmitter, and region of the central nervous system. We now propose to investigate the specific presynaptic actions and Na+ channel blocking mechanisms of inhaled anesthetics in detail to more fully understand how general anesthetics act so that future anesthetics can be developed with reduced undesirable side-effects and current anesthetics can be used more safely.